Factor the data into categorical variables
# Refactoring Columns for samples
data2$Sample_ID <- as.factor(data2$Sample_ID)
data2$Dilution_factor <- as.numeric(data2$Dilution_factor)
data2$Injection<- as.factor(data2$Injection)
data2$Tech_rep <- as.numeric(data2$Tech_rep)
data2
# Refactoring COlumns for key
key$Sample_ID <- as.factor(key$Sample_ID)
key$Animal <- as.factor(key$Animal)
key$Condition <- as.factor(key$Condition)
key
# Refactoring columns for standards
standards2$Sample_ID <- as.factor(standards2$Sample_ID)
standards2$When <- as.factor(standards2$When)
standards2$Dilution_factor <- as.numeric(standards2$Dilution_factor)
standards2$Injection <- as.factor(standards2$Injection)
standards2$Nano_day <- as.numeric(standards2$Nano_day)
standards2
Back calculate standards
standards2 <- standards2 %>%
mutate(True_Count=Dilution_factor*Count)
# Set the correct order of 'categorical factors'
standards2$Nano_day <- factor(standards2$Nano_day, levels=c('1'))
standards2$When <- factor(standards2$When, levels=c('before','after'))
standards2
Summarize three technical standard replicates
standards3 <- standards2 %>%
group_by(particle_size,Sample_ID,When,Dilution_factor,Nano_day,Injection) %>%
summarise( tech_N = length(True_Count),
tech_mean = mean(True_Count),
tech_sd = sd(True_Count),
tech_se = tech_sd/sqrt(tech_N))
standards3
Summarize standards by injection
standards4 <- standards3 %>%
group_by(Nano_day,When,particle_size) %>%
summarise( inj_N = length(tech_mean),
inj_mean = mean(tech_mean),
inj_sd = sd(tech_mean),
inj_se = inj_sd/sqrt(inj_N))
standards4
Plot before and after plots, facet by experimental day
std_plot <- standards4 %>%
ggplot(aes(x=particle_size,y=inj_mean,color=When))+
geom_ribbon(aes(ymin=inj_mean-inj_se, ymax=inj_mean+inj_se),alpha=0.2,fill = alpha('grey12', 0.2)) + #error bars
geom_line(size=2) + xlim(0,500)+ #line size, x-axis scale
scale_y_continuous(expand=c(0,0))+ #set bottom of graph
xlab("Particle Size") + # X axis label
ylab("\nMean Particle Concentration/ml\n") + # Y axis label
ggtitle("Nanosight Histogram of\nVirgin Mouse Plasma")+ #title
labs(color="Condition")+ #Label table title
facet_grid(. ~ When)
std_plot

Standards particle concentrations from each experimental day
standards_df <- standards4 %>%
group_by(Nano_day,When) %>%
summarise(total=sum(inj_mean))
standards_df
Bar graph of standards particle concentrations
standards_df %>%
ggplot(aes(x=Nano_day,y=total,fill=When))+
geom_col(position="dodge")+
scale_y_continuous(expand=c(0,0))+ #set bottom of graph
xlab("Experimental Day") + # X axis label
ylab("\nMean Particle Concentration/ml\n") + # Y axis label
ggtitle("Nanosight Histogram of\nVirgin Mouse Plasma")+ #title
labs(color="When") #Label table title

Intraassay variability
Intra.assay_cv <- standards_df %>%
group_by(Nano_day) %>%
summarise(Day_N = length(total),
Day_mean = mean(total),
Day_sd = sd(total),
Day_se = Day_sd/sqrt(Day_N),
Day_cv = Day_sd/Day_mean )
Intra.assay_cv